CN102268052A - Method for preparing camellianin A - Google Patents
Method for preparing camellianin A Download PDFInfo
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- CN102268052A CN102268052A CN2011102265889A CN201110226588A CN102268052A CN 102268052 A CN102268052 A CN 102268052A CN 2011102265889 A CN2011102265889 A CN 2011102265889A CN 201110226588 A CN201110226588 A CN 201110226588A CN 102268052 A CN102268052 A CN 102268052A
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Abstract
The invention provides a method for preparing camellianin A, which comprises: crushing Adinandra nitida Merr.ex H.L.Li. serving as a raw material, extracting in ethanol in a liquid dynamic continuous reflux extraction device, mixing extraction solution, recovering ethanol, purifying by using membrane separation, and loading filtrate on macroporous resin; and after absorption is finished, eluting with water and ethanol, collecting target fluid, concentrating, drying, obtaining a coarse product and separating the camellianin A from the coarse product by high speed countercurrent chromatography. The method is simple in process flow, high extraction efficiency, and good for industrial production; the preparation process is basically free from pollution and low in cost; and the purity of the prepared camellianin A is over 98 percent.
Description
Technical field
The invention belongs to the field of Chinese medicines, relate to the preparation method of a kind of camellin A, particularly relate to a kind of method of using high-speed countercurrent chromatography purifying camellin A.
Background technology
Adinandra nitida (
Adinandra nitidaMerr. ex Li), have another name called the red light wood of bright leaf, be commonly called as the clint dish,, mainly be distributed in areas such as China Guangxi, Guangdong and Guizhou, aboundresources for Theaceae Yang Tong platymiscium.Multiple efficacies such as Adinandra nitida has anti-inflammatory, detoxifies, brings down a fever, hemostasis and step-down among the peoplely are commonly used to prevent various inflammation, unknown tumour and wound etc., and effect is remarkable.In recent years, studies show that in the Adinandra nitida leaf that flavones content surpasses more than 30%, and effects such as significantly anti-oxidant, reducing blood-fat, antibiotic, hypertension are arranged.Wherein camellin A is a Flavonoid substances main in the Adinandra nitida leaf, and content can reach more than 25%.
Chinese patent (application number is 200810062110.5) " Preparation method and use of camellin A and camellin B ", this patent is a raw material with stone precipice tea, extract with the water-alcohol solvent system, concentrating under reduced pressure, with Semi-polarity organic solvent extracting extracting degreasing, water continues to concentrate shiyacha total flavone, total flavones is obtained camellin A through water alcohol system recrystallization, with total flavones salt of wormwood is to pass through cation-exchange chromatography or the pure system of direct water recrystallization after camellin A changes into camellin B, obtain camellin B, this method extraction yield is low, the sample loss amount is big, the cycle is long.
Yuan Erdong etc. (foodstuffs industry science and technology, 2008,4(29): 212-214) adopt the macroporous resin separation and purification to obtain 91.7% camellin A, the last tea glycosides A purity that this method obtained can not reach the requirement of reference substance.
Summary of the invention
The objective of the invention is provides the preparation method of a kind of camellin A in order to overcome the deficiencies in the prior art, uses high speed adverse current chromatogram separation and purification camellin A, simple to operate, sample loss is few, product purity is high.
To achieve these goals, technical scheme of the present invention is as follows:
The preparation method of a kind of camellin A is characterized in that concrete steps are as follows:
(1) extract: with the Adinandra nitida leaf is raw material, pulverizes, and places the dynamic continuous counter-flow extraction equipment alcohol extracting of liquid, serves as to extract solvent with the edible ethanol solution of 60-85%, under 40-65 ℃ extraction temperature, extracts 2-3 time united extraction liquid;
(2) membrane sepn: said extracted liquid reclaims ethanol, and the centrifuging and taking supernatant liquor carries out micro-filtration, ultrafiltration successively in the rolled film separating device, obtains seeing through liquid;
(3) column chromatography: see through macroporous resin on the liquid, after absorption was finished, water and 40-60% ethanol elution were collected ethanol eluate, and concentrate drying obtains camellin A crude product;
(4) high speed adverse current chromatogram separates: adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, prepare camellin A.
The dynamic continuous countercurrent device of liquid is that a plurality of extraction element serial or parallel connections or series-parallel connection are share in the described step (1), and extract the ethanolic soln consumption is 3-5 times (v/w) of raw materials quality at every turn.
Mould material is poly (ether sulfone) film or polysulfone membrane in the described step (2), and the microfiltration membrane aperture is 0.2-0.45 μ m, and the ultra-filtration membrane molecular weight cut-off is 30000-1000, and working pressure is 0.1-0.15Mpa.
Macroporous adsorbent resin in the described step (3) is AB-8, D3520, HPD100 or ADS-7 type.
Described step (4) high speed counter current chromatography separates solvent systems and selects normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution for use, volume ratio is (1-3): (3-5): (1-3): (4-6), standing demix behind the solvent systems mixing, below be stationary phase mutually, be moving phase mutually down, flow rate of mobile phase is controlled at 1.5-2.5ml/min, and engine speed is controlled at 750-900r/min.
The invention has the beneficial effects as follows:
(1) utilizes the dynamic Continuous Countercurrent Extraction technology of liquid, constantly extracts active ingredients is come out, shortened the consumption of extracting cycle and solvent greatly by the round-robin extracting solution.
(2) adopt membrane sepn, macroporous adsorbent resin and high-speed countercurrent chromatography to combine and separate preparation camellin A, technology is simple, cost is low, product content is high.
Further specify the present invention below in conjunction with embodiment, but the scope of protection of present invention is not limited to the following example.
Embodiment
Embodiment 1:
With the Adinandra nitida leaf is raw material, pulverize, get 3kg, on average place 3 placed in-line extraction elements, the edible ethanol solution that adds 12L60%, under 40 ℃ extraction temperature, extract each 2h 3 times, united extraction liquid, reclaim ethanol, the centrifuging and taking supernatant liquor carries out micro-filtration successively in the rolled film separating device, ultrafiltration, working pressure is 0.15Mpa, obtain seeing through liquid, by AB-8 type macroporous adsorptive resins, after absorption is finished, elder generation's water is eluted to colourless, use 50% ethanol elution again, collect ethanol eluate, concentrate drying obtains camellin A crude product, adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, be 1:3:1:4 preparation normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution solvent systems by volume, standing demix behind the solvent systems mixing, more than be stationary phase mutually, be moving phase mutually down, stationary phase is full of the high speed adverse current chromatogram post, opens engine speed and transfer to 800r/min, pump into down and make moving phase mutually, after the running balance, flow rate of mobile phase is controlled at 1.5ml/min, and by the sampling valve sample introduction, HPLC detected under crude product was dissolved in, collect target component, the dry flow point of concentrating under reduced pressure obtains camellin A, content 98.8%.
Embodiment 2:
With the Adinandra nitida leaf is raw material, pulverize, get 3kg, on average place 3 placed in-line extraction elements, the edible ethanol solution that adds 15L85%, under 40 ℃ extraction temperature, extract each 3h 2 times, united extraction liquid, reclaim ethanol, the centrifuging and taking supernatant liquor carries out micro-filtration successively in the rolled film separating device, ultrafiltration, working pressure is 0.15Mpa, obtain seeing through liquid, by HPD100 type macroporous adsorptive resins, after absorption is finished, elder generation's water is eluted to colourless, use 40% ethanol elution again, collect ethanol eluate, concentrate drying obtains camellin A crude product, adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, be 2:3:3:5 preparation normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution solvent systems by volume, standing demix behind the solvent systems mixing, more than be stationary phase mutually, be moving phase mutually down, stationary phase is full of the high speed adverse current chromatogram post, opens engine speed and transfer to 850r/min, pump into down and make moving phase mutually, after the running balance, flow rate of mobile phase is controlled at 1.5ml/min, and by the sampling valve sample introduction, HPLC detected under crude product was dissolved in, collect target component, the dry flow point of concentrating under reduced pressure obtains camellin A, content 98.5%.
Embodiment 3:
With the Adinandra nitida leaf is raw material, pulverize, get 5kg, on average place 5 placed in-line extraction elements, the edible ethanol solution that adds 25L80%, under 50 ℃ extraction temperature, extract each 4h 2 times, united extraction liquid, reclaim ethanol, the centrifuging and taking supernatant liquor carries out micro-filtration successively in the rolled film separating device, ultrafiltration, working pressure is 0.15Mpa, obtain seeing through liquid, by ADS-7 type macroporous adsorptive resins, after absorption is finished, elder generation's water is eluted to colourless, use 45% ethanol elution again, collect ethanol eluate, concentrate drying obtains camellin A crude product, adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, be 3:4:3:5 preparation normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution solvent systems by volume, standing demix behind the solvent systems mixing, more than be stationary phase mutually, be moving phase mutually down, stationary phase is full of the high speed adverse current chromatogram post, opens engine speed and transfer to 750r/min, pump into down and make moving phase mutually, after the running balance, flow rate of mobile phase is controlled at 2.5ml/min, and by the sampling valve sample introduction, HPLC detected under crude product was dissolved in, collect target component, the dry flow point of concentrating under reduced pressure obtains camellin A, content 98.6%.
Embodiment 4:
With the Adinandra nitida leaf is raw material, pulverize, get 5kg, on average place 5 placed in-line extraction elements, the edible ethanol solution that adds 25L70%, under 65 ℃ extraction temperature, extract each 3h 3 times, united extraction liquid, reclaim ethanol, the centrifuging and taking supernatant liquor carries out micro-filtration successively in the rolled film separating device, ultrafiltration, working pressure is 0.1Mpa, obtain seeing through liquid, by D3520 type macroporous adsorptive resins, after absorption is finished, elder generation's water is eluted to colourless, use 40% ethanol elution again, collect ethanol eluate, concentrate drying obtains camellin A crude product, adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, be 1:3:1:4 preparation normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution solvent systems by volume, standing demix behind the solvent systems mixing, more than be stationary phase mutually, be moving phase mutually down, stationary phase is full of the high speed adverse current chromatogram post, opens engine speed and transfer to 800r/min, pump into down and make moving phase mutually, after the running balance, flow rate of mobile phase is controlled at 2ml/min, and by the sampling valve sample introduction, HPLC detected under crude product was dissolved in, collect target component, the dry flow point of concentrating under reduced pressure obtains camellin A, content 98.1%.
Embodiment 5:
With the Adinandra nitida leaf is raw material, pulverize, get 5kg, on average place 5 placed in-line extraction elements, the edible ethanol solution that adds 25L80%, under 55 ℃ extraction temperature, extract each 4h 2 times, united extraction liquid, reclaim ethanol, the centrifuging and taking supernatant liquor carries out micro-filtration successively in the rolled film separating device, ultrafiltration, working pressure is 0.1Mpa, obtain seeing through liquid, by ADS-7 type macroporous adsorptive resins, after absorption is finished, elder generation's water is eluted to colourless, use 60% ethanol elution again, collect ethanol eluate, concentrate drying obtains camellin A crude product, adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, be 2:5:2:4 preparation normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution solvent systems by volume, standing demix behind the solvent systems mixing, more than be stationary phase mutually, be moving phase mutually down, stationary phase is full of the high speed adverse current chromatogram post, opens engine speed and transfer to 900r/min, pump into down and make moving phase mutually, after the running balance, flow rate of mobile phase is controlled at 2ml/min, and by the sampling valve sample introduction, HPLC detected under crude product was dissolved in, collect target component, the dry flow point of concentrating under reduced pressure obtains camellin A, content 98.2%.
Embodiment 6:
With the Adinandra nitida leaf is raw material, pulverize, get 5kg, on average place 5 placed in-line extraction elements, the edible ethanol solution that adds 20L60%, under 65 ℃ extraction temperature, extract each 4h 2 times, united extraction liquid, reclaim ethanol, the centrifuging and taking supernatant liquor carries out micro-filtration successively in the rolled film separating device, ultrafiltration, working pressure is 0.15Mpa, obtain seeing through liquid, by HPD100 type macroporous adsorptive resins, after absorption is finished, elder generation's water is eluted to colourless, use 60% ethanol elution again, collect ethanol eluate, concentrate drying obtains camellin A crude product, adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, be 3:5:3:6 preparation normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution solvent systems by volume, standing demix behind the solvent systems mixing, more than be stationary phase mutually, be moving phase mutually down, stationary phase is full of the high speed adverse current chromatogram post, opens engine speed and transfer to 850r/min, pump into down and make moving phase mutually, after the running balance, flow rate of mobile phase is controlled at 2.5ml/min, and by the sampling valve sample introduction, HPLC detected under crude product was dissolved in, collect target component, the dry flow point of concentrating under reduced pressure obtains camellin A, content 98.3%.
Claims (5)
1. the preparation method of a camellin A is characterized in that concrete steps are as follows:
(1) extract: with the Adinandra nitida leaf is raw material, pulverizes, and places the dynamic continuous counter-flow extraction equipment alcohol extracting of liquid, serves as to extract solvent with the edible ethanol solution of 60-85%, under 40-65 ℃ extraction temperature, extracts 2-3 time united extraction liquid;
(2) membrane sepn: said extracted liquid reclaims ethanol, and the centrifuging and taking supernatant liquor carries out micro-filtration, ultrafiltration successively in the rolled film separating device, obtains seeing through liquid;
(3) column chromatography: see through macroporous resin on the liquid, after absorption was finished, water and 40-60% ethanol elution were collected ethanol eluate, and concentrate drying obtains camellin A crude product;
(4) high speed adverse current chromatogram separates: adopt high-speed countercurrent chromatography that the gained crude product is carried out separation and purification, prepare camellin A.
2. the preparation method of a kind of camellin A according to claim 1, it is characterized in that: the dynamic continuous countercurrent device of liquid is that a plurality of extraction element serial or parallel connections or series-parallel connection are share in the described step (1), and extract the ethanolic soln consumption is 3-5 times (v/w) of raw materials quality at every turn.
3. the preparation method of a kind of camellin A according to claim 1, it is characterized in that: mould material is poly (ether sulfone) film or polysulfone membrane in the described step (2), the microfiltration membrane aperture is 0.2-0.45 μ m, and the ultra-filtration membrane molecular weight cut-off is 30000-1000, and working pressure is 0.1-0.15Mpa.
4. the preparation method of a kind of camellin A according to claim 1 is characterized in that: in the described step (3)
Macroporous adsorbent resin be AB-8, D3520, HPD100 or ADS-7 type.
5. the preparation method of a kind of camellin A according to claim 1, it is characterized in that: described step (4) high speed counter current chromatography separates solvent systems and selects normal hexane-propyl carbinol-methyl alcohol-0.5% acetic acid aqueous solution for use, volume ratio is (1-3): (3-5): (1-3): (4-6), solvent systems preparation back standing demix, below be stationary phase mutually, be moving phase mutually down, flow rate of mobile phase is controlled at 1.5-2.5ml/min, and engine speed is controlled at 750-900r/min.
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| CN2011102265889A CN102268052A (en) | 2011-08-09 | 2011-08-09 | Method for preparing camellianin A |
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| CN2011102265889A CN102268052A (en) | 2011-08-09 | 2011-08-09 | Method for preparing camellianin A |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102628839A (en) * | 2012-02-22 | 2012-08-08 | 广西壮族自治区中医药研究院 | Preparation of high-purity camellianin A and its quality control method |
| CN104998263A (en) * | 2015-09-02 | 2015-10-28 | 集美大学 | Method for quickly separating alpha-glucosidase inhibitor from camellia pollen |
| CN111297979A (en) * | 2020-04-10 | 2020-06-19 | 华南理工大学 | Application of Shibi tea and/or camellin A in preparation of medicine for treating gastric ulcer |
-
2011
- 2011-08-09 CN CN2011102265889A patent/CN102268052A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102628839A (en) * | 2012-02-22 | 2012-08-08 | 广西壮族自治区中医药研究院 | Preparation of high-purity camellianin A and its quality control method |
| CN102628839B (en) * | 2012-02-22 | 2013-10-30 | 广西壮族自治区中医药研究院 | Preparation of high-purity camellianin and its quality control method |
| CN104998263A (en) * | 2015-09-02 | 2015-10-28 | 集美大学 | Method for quickly separating alpha-glucosidase inhibitor from camellia pollen |
| CN104998263B (en) * | 2015-09-02 | 2017-11-03 | 集美大学 | A kind of method of the quick separating α glucosidase inhibitors from camellia pollen |
| CN111297979A (en) * | 2020-04-10 | 2020-06-19 | 华南理工大学 | Application of Shibi tea and/or camellin A in preparation of medicine for treating gastric ulcer |
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Application publication date: 20111207 |